In this project, understanding of the structure/function relationships for the major RNA's in translation is sought. The method to be used is site-directed mutagenesis of tRNA, mRNA, or rRNA genes, selection or screening of function, then quantitative comparison of action of altered RNA products in vivo. There are three major divisions to the work. 1. The extended anticodon hypothesis. In the first, tRNA genes will be manipulated to show that a second class of anticodons require an optimal anticodon arm structure for translational efficiency, as do Uxx codons. The Uxx series has been investigated in the preceding grant period. Ribosomal RNA:tRNA interaction, suggested by these prior results, will be tested by genetic means. 2. The A24 mutation. In the second, an important tRNA mutant which seems to be an exception to the generalization of the first section, will be investigated to see if the hypothesis needs to be generalized to embrace it. 3. Codon choice and codon context. In the third section of the grant, both messages and tRNA's will be altered to investigate the differential effects of codon choice and codon context on gene expression. Message: tRNA interactions and tRNA:tRNA interactions will be investigated, and a new theory will be tested to connect the local rate of translation to gene expression.